Abstract
Purpose :
Cornea alkali burn injuries are typically extremely serious, difficult to treat, potentially blinding and can lead to loss of the affected eye. Although studied for many years, there are no alkali wound-specific treatments available. The CYP450 arachidonic acid metabolism pathway generates epoxide fatty acids (EpFAs) which have analgesic, inflammation-resolving and anti-fibrotic activities in several tissues, but their influence on cornea anti-fibrotic activity has never been examined. Our studies were designed to examine the role of soluble epoxide hydrolase (sEH), which metabolizes EpFAs, and EpFAs, in alkali-wounded corneas.
Methods :
Cornea alkali injuries were created in C57BL/6J mice with 0.25N NaOH or 20% ammonia (AMM). The EpFAs 17,18-epoxyeicosatetraenoic acid (EEQ) and epoxyeicosatrienoic acid (mixture of regioisomers; EET), a biostable (n-propoxy) EET mimic, an epoxydocosapentaenoic acid mimic (MBD-139-5 EpDPA) , and the sEH inhibitor TPPU, were topically applied to eyes for up to 17 days post-wounding. The degree of corneal opacification and neovascularization were scored in a blinded fashion. Cell culture experiments were conducted with mouse cornea fibroblasts (MSC) briefly being exposed to 1% AMM in DMEM and treated with EpFAs. Human cornea fibroblasts (HSC) were cultured with 10 ng/ml TGFβ1 and EpFAs for 48 hours. Western blotting and immnostaining were used to detect TGFβα-induced α-smooth muscle actin (α-SMA), collagen III, and sEH expression.
Results :
NaOH wounded eyes developed haziness and fibrosis, with AMM wounded eyes becoming fibrotic with neovascularization. Haziness/fibrosis was dramatically reduced or eliminated in both NaOH and AMM wounded mouse corneas treated with EpFAs and sEHi by day 17. EpFA and sEHi treated AMM corneas did not vascularize. α-SMA and collagen III protein levels was significantly decreased in NaOH wounded mouse corneas treated with EEQ, and decreased in AMM wounded corneas treated with TPPU. MSC exposed to AMM had altered phenotypes that were rescued following EpFA treatment. TGF β1-stimulated α-SMA, collagen III, and sEH protein levels were significantly reduced in HSC treated with EpFAs and sEHi.
Conclusions :
EpFAs play a critical role in NaOH and AMM cornea injury responses as demonstrated by in vivo animal models and in vitro cell culture methods. Topical sEHi and/or EpFA administration can effectively treat these wounds.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.